CD8+ cytotoxic Tcells (CTLs) are important effector cells in antiviral and anticancer immunity, and these cells are promising candidates for adoptive immunotherapy of cancer. The exquisite specificity toward antigen-bearing cells (cognate targets) is one of the most important properties of CD8+ CTL-mediated cytotoxicity. However, we found that in the presence of antigen-bearing targets the CTLs acquire the ability to efficiently lyse noncognate target cells ("innocent bystanders") by a Fas ligand (FasL)/Fas-based cytotoxicity mechanism. Unlike anti-Fas mAb-induced cell death, the lysis of bystanders was not only FasL/Fas dependent but also required the adhesion molecule LFA-1 on the surface of the activated CTL and the presence of ICAM-1 molecules on bystanders. According to our model, the encounter with the antigen-bearing target results in the Tcell antigen receptor (TCR)-induced upregulation of FasL and in activation of cell adhesion LFA-1 molecules on the CTL surface. This enables CTLs to form "productive" conjugates and to kill those bystanders that express both functional Fas molecules and the LFA-1 counter receptor, ICAM-1. Fas-expressing bystanders are vulnerable only if they are encountered no later than 2h after the latest conjugation of CTL with the antigen-bearing targets. The lysis of bystanders is minimized by the activity of constitutively active and cell surface membrane-associated PP2a protein phosphatase in CTL. The TCR is not the only activator of FasL-mediated cytotoxicity, since Thy-1 surface antigens are also able to induce the lysis of Fas-expressing targets by different CTL lines, clones, or T-helper hybridoma cells. The use of TCR-CD3 mutants demonstrated that Thy-1-mediated upregulation of FasL in T-helper hybridoma cells requires an intact TCR-coupled intracellular pathway. The ability of Thy-1 surface proteins to upregulate FasL points to the potential FasL-upregulating properties of other GPI-linked Tcell surface proteins. We propose that the lysis of bystanders represents an acceptable "collateral" damage to Fas-expressing bystanders during normal immune response, but the persistent presence of activated CTLs could result in immunopathologies involving such tissues as heart and liver.